Abstract:

Real-time quantitative PCR is often used to analyze the target gene expression variation. The most commonly used methods to analyze data from real-time quantitative PCR include two types: absolute quantification and relative quantification. To avoid the complex of absolute quantification and simplify the relative quantification, the methods of 2^-ΔΔCT, 2^-ΔCT and standard curve were used to analyze the OsRDB1 expression variation under different chemical and hormone treatments in this study. The ratio of gene expression variation was calculated with the slope of standard curve. The results revealed that the 2^-ΔΔCT method had to be introduced a housekeep gene as a control, with a complicated calculation and the relative ratio was deviated by PCR amplification efficiency. The 2^-ΔCT and standard curve methods were convenient and saved chemicals used in the quantitative PCR, but the results were liable to the effect of the veracity of the RNA concentration. The modified method of standard curve introduced the curve slope to eliminate the effect of amplification efficiency, and the result of standard curve method most closely revealed the diversity of original copy.

Key words: Relative quantification, Standard curve, Real-time quantitative PCR, OsRDB1